We propose to isolate and purify plasma membrane tumor-associated antigen(s) (TAA) of human osteosarcoma from biopsied and autopsied tumor tissues and from osteosarcoma cell lines. Plasma membrane TAA will be isolated using various solubilization technique (papain, 6 M guanidine HC1, 0.5% NP40, and sonication). Antigenically active isolated plasma membrane TAA will be further purified by a combination of methods, including DEAE anion-exchange chromatography, Sephadex G-200 gel filtration, preparative polyacrylamide gel electrophoresis, and affinity chromatography. Purified plasma membrane TAA will be analyzed by analytical gel electrophoresis, and isoelectric focusing. Antigenic activities of the isolated and purified plasma membrane TAA will be determined by complement fixation assay, lymphocyte stimulation assay, blocking assay, leukocyte migration inhibition assay, and skin testing of osteosarcoma patients immunodiffusion, and co-precipitation assays. Hybridomas will be made to produce monoclonal antibodies for osteosarcoma associated antigens when active plasma membrane TAA of adequate purity has been obtained. A radioimmunoassay (RIA) will be developed using the monoclonal antibodies for the determination of antigen levels in the sera of osteosarcoma patients. The clinical applications of the RIA will be examined by comparing antigen levels in the sera of osteosarcoma patients at different stages of disease (including those with and without metastases and those in remission), patients with non-neoplastic bone diseases, and normal individuals.